Growth cones are highly specialized structures located at the tips of axons that integrate a complex concert of chemical cues to guide axons to their appropriate destinations. This extraordinary navigational ability of growth cones is essential for proper formation of the nervous system. Aberrant axonal projections are thought to contribute to conditions such as mental retardation. Furthermore, the failure of damaged adult axons to re-navigate to their targets after spinal cord injury or stroke contributes to paralysis and/or compromised motor, sensory and cognitive function. Therefore, a better understanding of growth cone navigation will contribute to treatment of these conditions. For this reason, it is essential to understand the extracellular ligands, as well as their receptors, that direct growth cone guidance. While growth cone ligand-receptor complexes are beginning to be unraveled, significant gaps in our knowledge still exist. In this proposal, we investigate the relationship between the axon guidance molecule netrin-1 and integrin receptors in chick sensory neurons. While netrin-1 has many known receptors, these studies are the first to highlight integrins as possible netrin-1 receptors and as mediators of netrin-induced growth cone behaviors, such as collapse. Our preliminary data suggest that two integrin subunits ( 3, 6) are involved in netrin-induced growth cone collapse. Moreover, our data show that netrin-1 causes integrin activation (a conversion to a high ligand affinity conformational state). These data are exciting because they suggest that integrin receptors, well known for their ability to interact with extracellular matrix molecules, may also interact with the axon guidance molecule, netrin-1 and play a role in netrin-mediated growth cone behaviors. In Aim 1 we will we use siRNA, function blocking antibodies and competitive peptides to individually test the involvement of all known integrin subunits on netrin-induced growth cone collapse. These experiments will provide a profile of neuronal integrin subunits required for netrin- mediated collapse. In Aims 2 and 3, we will examine if the integrin-netrin interaction is direct and/or indirect. In Aim 2, we will use co-immunoprecipitations and affinity columns to determine if netrin-1 directly binds to integrin receptors. This aim will determine if integrins may serve as netrin receptors. In Aim 3, we will determine if traditional netrin receptors mediate an integrin-netrin interaction. More specifically, we will use siRNA to investigate if traditional netrin receptors are involved in netrin-induced integrin activation. These studies will reveal an indirect integrin-netrin relationship. Aims 2 and 3 are not mutually exclusive;netrin-1 may interact with integrins in both a direct (Aim 2) and indirect (Aim 3) manner. This research will provide an excellent training opportunity for undergraduates as they acquire the skills of scientific thinking and engage in all aspects of scientific research. Furthermore, these studies will establish the relationship between netrin-1 and integrin receptors as well as the impact of this relationship on growth cone guidance. These results will provide new insights into the role of integrin receptors in netrin-mediated growth cone repulsive behaviors. PUBLIC HEALTH RELEVANCE: The ability of millions of neurons to accurately navigate their axonal projections through a complex extracellular environment to reach their appropriate targets is essential for proper formation of the nervous system and yet the mechanisms of this navigation are not fully understood. We propose to investigate the role of cell adhesion molecules, integrin receptors, in the navigational abilities of neurons in response to an extracellular guidance molecule, netrin-1.These studies will provide fundamental insights into the mechanisms of axonal guidance and are expected to ultimately lead to the development of new therapies for the treatments of spinal cord injuries or disorders arising from deficiencies in this process.